Real-time label-free detection of dynamic aptamer-small molecule interactions using a nanopore nucleic acid conformational sensor.

Nucleic acids can undergo conformational changes upon binding small molecules. These conformational changes can be exploited to develop new therapeutic strategies through control of gene expression or triggering of cellular responses and can also be used to develop sensors for small molecules such as neurotransmitters. Many analytical approaches can detect dynamic conformational change of nucleic acids, but they need labeling, are expensive, and have limited time resolution. The nanopore approach can provide a conformational snapshot for each nucleic acid molecule detected, but has not been reported to detect dynamic nucleic acid conformational change in response to small -molecule binding. Here we demonstrate a modular, label-free, nucleic acid-docked nanopore capable of revealing time-resolved, small molecule-induced, single nucleic acid molecule conformational transitions with millisecond resolution. By using the dopamine-, serotonin-, and theophylline-binding aptamers as testbeds, we found that these nucleic acids scaffolds can be noncovalently docked inside the MspA protein pore by a cluster of site-specific charged residues. This docking mechanism enables the ion current through the pore to characteristically vary as the aptamer undergoes conformational changes, resulting in a sequence of current fluctuations that report binding and release of single ligand molecules from the aptamer. This nanopore tool can quantify specific ligands such as neurotransmitters, elucidate nucleic acid-ligand interactions, and pinpoint the nucleic acid motifs for ligand binding, showing the potential for small molecule biosensing, drug discovery assayed via RNA and DNA conformational changes, and the design of artificial riboswitch effectors in synthetic biology.

Autophagic UVRAG Promotes UV-Induced Photolesion Repair by Activation of the CRL4(DDB2) E3 Ligase.

UV-induced DNA damage, a major risk factor for skin cancers, is primarily repaired by nucleotide excision repair (NER). UV radiation resistance-associated gene (UVRAG) is a tumor suppressor involved in autophagy. It was initially isolated as a cDNA partially complementing UV sensitivity in xeroderma pigmentosum (XP), but this was not explored further. Here we show that UVRAG plays an integral role in UV-induced DNA damage repair. It localizes to photolesions and associates with DDB1 to promote the assembly and activity of the DDB2-DDB1-Cul4A-Roc1 (CRL4(DDB2)) ubiquitin ligase complex, leading to efficient XPC recruitment and global genomic NER. UVRAG depletion decreased substrate handover to XPC and conferred UV-damage hypersensitivity. We confirmed the importance of UVRAG for UV-damage tolerance using a Drosophila model. Furthermore, increased UV-signature mutations in melanoma correlate with reduced expression of UVRAG. Our results identify UVRAG as a regulator of CRL4(DDB2)-mediated NER and suggest that its expression levels may influence melanoma predisposition.

Teaching antenatal hand expression: a feasibility study in an inner urban U.S. hospital.

BACKGROUND: Many women have low confidence in breastfeeding and have concerns regarding low milk volume or discomfort with breastfeeding. Antenatal hand expression may be an opportunity to help women feel more comfortable with breastfeeding and help promote exclusive breastfeeding. A study at a hospital in Philadelphia, Pennsylvania, U.S. assessed the feasibility of teaching antenatal hand expression at 39 weeks among socioeconomically disadvantaged populations, overall participant satisfaction and adoption of hand expression and breastfeeding. METHODS: From March 2020 to June 2021, women recruited at 34-39 weeks were taught to hand express, collect, and store colostrum. Starting from 39 weeks, participants were asked to practice hand expression 1-3 times / day until delivery, log their experiences, and store colostrum expressed. Women were contacted to encourage continued hand expression and answer any questions. Postpartum, a survey assessed satisfaction with hand expression and issues encountered. The survey also inquired about breastfeeding plans and barriers, and whether women were exclusively breastfeeding (defined as infants who received only breastmilk from the time of birth). Chart review of postpartum or well-baby visit notes determined whether women continued breastfeeding. RESULTS: Of the 29 participants, 72% (21/29) reported hand expressing at home, and no women reported contractions when hand expressing. Participants rated mean satisfaction of 8.1/10 (SD = 1.62) with antenatal hand expression, mean satisfaction of 9.4/10 (SD = 0.90) toward hand expression education, mean likelihood of 9.4/10 (SD = 1.24) recommending hand expression to others, and a mean score of 8.1/10 (SD = 1.69) on how helpful hand expression was in breastfeeding initiation. 90% (26/29) of women initiated breastfeeding after birth and 72% (21/29) exclusively breastfed on discharge, but only 11/29 (38%) continued exclusively breastfeeding when re-assessed 4-6 weeks postpartum. Barriers included maternal discomfort, low milk supply, and maternal or infant illness. CONCLUSIONS: This study suggests that women in an urban setting would be willing to practice antenatal hand expression. A larger and adequately powered study could be feasible to determine associations between antenatal hand expression and breastfeeding rates and confidence.

Machine learning and image analysis in vascular surgery.

Deep learning, a subset of machine learning within artificial intelligence, has been successful in medical image analysis in vascular surgery. Unlike traditional computer-based segmentation methods that manually extract features from input images, deep learning methods learn image features and classify data without making prior assumptions. Convolutional neural networks, the main type of deep learning for computer vision processing, are neural networks with multilevel architecture and weighted connections between nodes that can "auto-learn" through repeated exposure to training data without manual input or supervision. These networks have numerous applications in vascular surgery imaging analysis, particularly in disease classification, object identification, semantic segmentation, and instance segmentation. The purpose of this review article was to review the relevant concepts of machine learning image analysis and its application to the field of vascular surgery.

Childhood pneumonia in humanitarian emergencies in low- and middle-income countries: A systematic scoping review.

Background: Humanitarian emergencies increase many risk factors for pneumonia, including disruption to food, water and sanitation, and basic health services. This review describes pneumonia morbidity and mortality among children and adolescents affected by humanitarian emergencies. Methods: We searched MEDLINE, EMBASE, and PubMed databases for publications reporting pneumonia morbidity or mortality among children aged 1 month to 17 years in humanitarian emergencies (eg, natural disaster, armed conflict, displacement) in low- and middle-income countries (LMICs). Results: We included 22 papers published between January 2000 and July 2021 from 33 countries, involving refugee/displaced persons camps (n = 5), other conflict settings (n = 14), and natural disaster (n = 3). Population pneumonia incidence was high for children under 5 years of age (73 to 146 episodes per 100 patient-years); 6%-29% met World Health Organization (WHO) criteria for severe pneumonia requiring admission. Pneumonia accounted for 13%-34% of child and adolescent presentations to camp health facilities, 7%-48% of presentations and admissions to health facilities in other conflict settings, and 12%-22% of admissions to hospitals following natural disasters. Pneumonia related deaths accounted for 7%-30% of child and adolescent deaths in hospital, though case-fatality rates varied greatly (0.5%-17.2%). The risk for pneumonia was greater for children who are: recently displaced, living in crowded settings (particularly large camps), with deficient water and sanitation facilities, and those who are malnourished. Conclusion: Pneumonia is a leading cause of morbidity and mortality in children and adolescents affected by humanitarian emergencies. Future research should address population-based pneumonia burden, particularly for older children and adolescents, and describe contextual factors to allow for more meaningful interpretation and guide interventions.

A single centre audit: repeat pre-operative colonoscopy.

BACKGROUND: Repeat colonoscopy may be required for tumour localisation. The aim of the study is to explore the clinical settings it was used and benchmark the quality of initial colonoscopy against standardized guidelines for tumour localisation, tattooing and colonoscopy reporting amongst clinicians. METHODS: A retrospective study from 2016 to 2021 has been performed on patients who underwent elective colorectal cancer resections at the Northern Hospital. Patient demographics, colonoscopic and operative details were retrieved from the Bi-National Colorectal Cancer Audit (BCCA) Registry database and hospital medical records. PRIMARY OUTCOMES: changes in operative approach and delays to operation. SECONDARY OUTCOMES: reasons for a repeat colonoscopy and complications from repeat colonoscopy. RESULTS: A total of 339 patients were included in this study. 94 (28.6%) underwent a repeat colonoscopy. Re-scoping rate was 29.6% for surgeons, and 26.2% for non-operating endoscopists. Surgeons had a 5.9% localisation error rate, and non-operating endoscopist 6.95% (p = 0.673). Surgeons did not have a lower rate of repeat colonoscopy (p = 0.462). Repeat endoscopy was associated with a longer time to definitive operation (p < 0.001). No complications were associated with a repeat colonoscopy. CONCLUSION: There was no difference in localisation error rates or repeat colonoscopy amongst surgeons (29.6%) and non-operating endoscopists (26.2%) (p = 0.462). This could be explained by the standardized endoscopy training in Australia governed by a common training board. Lack of tattooing at index colonoscopy and inadequate documentation often led to a repeat endoscopy, which was associated with a longer time to definitive operation. Standardized guidelines in tattooing of lesions and colonoscopy reporting should be implemented.

Durational patterning at syntactic and discourse boundaries in Mandarin spontaneous speech.

This study focused on durational cues (i.e., syllable duration, pause duration, and syllable onset intervals (SOIs)) at discourse boundaries in two dialects of Mandarin,Taiwan and Mainland varieties. Speech was elicited by having 18 participants describe events in The Pear Story film. Recorded data were transcribed, labeled, and segmented into clauses. Discourse boundary indices were used to label discourse disjuncture levels. Results showed that the scope of lengthening included both the final and the penultimate syllables. Pause was a robust but optional indicator for discourse boundaries, and Taiwan Mandarin preferred unfilled over filled ones, while the Mainland variety did not show such strong preferences. Discourse hierarchy corresponded consistently with occurrences of pauses and duration of SOIs, within which pause was the main contributor. Higher discourse levels were more likely to be accompanied by pauses, and were indicated by longer pauses and SOIs. Syllable duration only played a secondary role in indicating discourse disjuncture size when pause was absent.When there was an accompanying pause, Mainland Mandarin relied solely on it to indicate discourse hierarchy, while Taiwan Mandarin used both syllable and pause duration, a dialectal difference that seemed to be rhythm-related. By shortening the degree of lengthening in the boundary syllable and lengthening the following pause at the same time,Taiwan Mandarin increased the absolute and relative duration of pause and maximized its role in indicating discourse hierarchy.The results of this study implied that the use of pause in discourse disjuncture demarcation was more of a language-specific choice while its role in discourse hierarchy encoding is more language-universal.

Ethanol Concentration of Kombucha Teas in British Columbia, Canada.

ABSTRACT: Kombucha is a sweetened tea beverage fermented by bacterial and yeast cultures. Sweeteners, such as glucose, sucrose, fructose, and others are converted by yeasts into ethanol and then by Acetobacter and other bacterial species into a weak acetic acid solution that is diluted, flavored, and packaged into glass or aluminum cans for consumer consumption. Naturally, fermented kombucha contains 0 to 3% alcohol by volume (ABV). However, kombucha containing ethanol is concerning for pregnant women and young children for whom low levels of ethanol consumption (<3% ABV) create adverse medical outcomes. In the province of British Columbia (BC), Canadian beverages containing >1% ABV are regulated as liquor. This study assessed ethanol concentrations in kombucha collected from processors and purchased at retail venues in BC. Ethanol values were compared with the place of manufacture (country or province) and place of purchase (grocery stores, restaurants, farmers' markets, recreational centers, and processors). Ethanol (n = 684) levels were measured by using a headspace gas chromatography-mass spectrometry method with a detection limit of 0.0002% ABV for ethanol. Overall, teas contained mean and median ethanol of 0.77 and 0.62% ABV, respectively, ranging from nondetectable up to 3.62% ABV. Four kombucha teas (0.6%) made by BC processors tested over 3% ABV, and 31.5% of samples contained ethanol that exceeded the BC regulatory limits for nonalcoholic beverages of 1% ABV. Kombucha manufactured in BC had significantly higher mean ethanol values (1.16% ABV) in comparison to all other places of manufacture. Similarly, mean ethanol tea values obtained from BC processors (1.2% ABV) and restaurants (1.01% ABV) were significantly higher than those obtained at other retail venues. This study demonstrates the potential for alcohol harm to at-risk populations consuming kombucha teas sold in BC.

Severe fever with thrombocytopenia syndrome phlebovirus non-structural protein activates TPL2 signalling pathway for viral immunopathogenesis.

Severe fever with thrombocytopenia syndrome phlebovirus (SFTSV), listed in the World Health Organization Prioritized Pathogens, is an emerging phlebovirus with a high fatality1-4. Owing to the lack of therapies and vaccines5,6, there is a pressing need to understand SFTSV pathogenesis. SFSTV non-structural protein (NSs) has been shown to block type I interferon induction7-11 and facilitate disease progression12,13. Here, we report that SFTSV-NSs targets the tumour progression locus 2 (TPL2)-A20-binding inhibitor of NF-κB activation 2 (ABIN2)-p105 complex to induce the expression of interleukin-10 (IL-10) for viral pathogenesis. Using a combination of reverse genetics, a TPL2 kinase inhibitor and Tpl2-/- mice showed that NSs interacted with ABIN2 and promoted TPL2 complex formation and signalling activity, resulting in the marked upregulation of Il10 expression. Whereas SFTSV infection of wild-type mice led to rapid weight loss and death, Tpl2-/- mice or Il10-/- mice survived an infection. Furthermore, SFTSV-NSs P102A and SFTSV-NSs K211R that lost the ability to induce TPL2 signalling and IL-10 production showed drastically reduced pathogenesis. Remarkably, the exogenous administration of recombinant IL-10 effectively rescued the attenuated pathogenic activity of SFTSV-NSs P102A, resulting in a lethal infection. Our study demonstrates that SFTSV-NSs targets the TPL2 signalling pathway to induce immune-suppressive IL-10 cytokine production as a means to dampen the host defence and promote viral pathogenesis.